Glaucoma is one of the leading causes of blindness throughout the world, but the cellular and molecular mechanisms are mostly unknown. Insights into this disease have come from genetic studies in animal models and humans. One point mutation in the gene optineurin (optn), E50K, has been implicated in causing disease in as many as 13.5% of primary open angle glaucoma cases for one population of glaucoma patients. Through in vitro studies, optn function has been implicated in two diverse cellular processes: NF-?B signaling and vesicle trafficking. However, it is not known whether wild-type optn or the E50K mutation alters these pathways in vivo and specifically within the eye. Furthermore, which cell types are responsible for generating glaucoma-associated pathology when optn is mutated remains a mystery. Therefore, this proposal will conduct a broad survey of these cellular processes to determine whether optn or E50K can alter their function in retinal cell types. This will be accomplished by generating zebrafish transgenic lines that express wild-type optn or the E50K variant (E32K in zebrafish). First, the cell autonomy of optn or E32K to cause glaucoma- associated pathology will be examined by comparing the phenotype when these cells are expressed ubiquitously versus expressed within retinal ganglion cells or glia, two cell types that are highly relevant to glaucoma pathogenesis. Second, the ability of these proteins to inhibit NF-?B signaling in the vertebrate eye will be tested, as well as how this inhibition may be contributing to glaucoma pathologies. Third, optn or E32K will be examined for their ability to alter vesicle trafficking of glutamate receptors to alter calcium dynamics in retinal ganglion cells, possibly leading to excitotoxicity. This work should provide an understanding of the cellular processes that optn affects in those cell types. This research will also determine potential mechanisms by which the E50K mutation can cause glaucoma-associated pathology. PUBLIC HEALTH RELEVANCE: Glaucoma is one of the leading causes of blindness throughout the world. Optineurin is a gene that has been implicated in causing a subset of glaucomas, but its functions within specific cell types, and how its dysfunction results in glaucoma-associated pathology are not known. In this project, I propose to study the function of both wild-type optineurin and a common mutant variant to determine in which signaling pathways these variants are acting, and in which cells they act to cause glaucoma- associated pathology.